Using Fractogel® EMD Tentacle Supports. Ion Exchange Chromatography . 3: BSA binding capacity of Fractogel® EMD TMAE (S) at linear flow rates up to . Fractogel® EMD TMAE Hicap (M). Ion Exchange chromatography using strong anion exchangers. Fractogel® ion exchangers are cross-linked. Sigma-Aldrich offers EMD Millipore, MiniChrom Column Fractogel® TMAE (S), 1 ml for your research needs. Find product specific information including.
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A miniaturized flowcell was fabricated similar in design ffractogel that used by Hubbuch and Kula A thin layer of resin slurry was pipetted onto a glass slide which had been pre-coated in gold and mounted onto a copper block. SEM and batch uptake experiments are used to give initial indications of foulant location and resin performance as fouling progresses, before CLSM is used to conduct a more detailed investigation.
A detailed account has been presented in the literature Iskra et al. The shape of the breakthrough went from sharp to diffuse as fouling progressed Fig. Confocal laser scanning microscopy CLSM during live uptake.
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Generation of Fouled Resin Samples Three resin samples were used in subsequent experimental studies to characterize the fouling. Experimental methodology for investigating clean, partially found, and extensively fouled resin samples. Visualising intraparticle protein transport in porous adsorbents by confocal microscopy. Bound BSA concentration as a function of time. Fouling on partially fouled resin surface is not as obvious, but does show an intermediate level of pore blockage Fig.
The results suggest that progressive fouling of resin can severely impact the performance of chromatography columns. In this article, scanning electron microscopy SEMbatch uptake experiments, confocal laser scanning microscopy CLSM and small-scale column studies were applied to characterize a case study where fouling had been observed during process development.
Cation exchange chromatography of monoclonal antibodies: The increased understanding into the nature of the fouling can help in the continued process development of this industrial example.
Integrating the area ftactogel the radial light intensity profiles, and correcting for the spherical nature of resin particles, indicates the relative amount of BSA bound to the different resin samples throughout uptake Fig. Downstream processing prior to the AEX step considered in this work consisted of centrifugation and depth filtration, followed by protein A chromatography. However, the amount of BSA bound to the partially and extensively fouled resin samples was approaching that of the clean resin at the end of the experiment.
Support Center Support Tmze. The use of BSA for breakthrough studies was not intended to replicate industrial process behavior. Processing of cell homogenates.
Great care was taken to ensure that the resin was not over or under packed. The results indicated that the foulant has a significant impact on the kinetics of adsorption, severely decreasing the protein uptake rate, but only results in a minimal decrease in saturation capacity.
In this article, we aim to elucidate on this resin fouling case study, by revealing the location of the foulant, and determining the mechanistic effects fouling has on protein uptake kinetics ffractogel resin capacity. The control experiment using clean resin at 0. The AEX columns used in this study were 0.
We utilized ImageJ v1.
In Figure 4images CX and CY illustrate the magnitude of the fouling, with many particles showing completely clogged pore entrances over a significant percentage of particle surface area. The overall objective of this work was to determine the location of fouling on resin particles and the effect of this rractogel on protein kinetics and resin capacity in an AEX polishing step from an industrial purification process.
Discoloration of ceramic hydroxyapatite used for protein chromatography. Live Uptake Data Processing The large number of confocal images from the flowcell experiments were processed to generate a reliable set of radial light intensity profiles.
In fractotel, when the resin was extensively fouled, onset of breakthrough was rapid, beginning after less than 1 CV, similar to what would be expected during operation in flow through mode with mtae protein binding. All chemicals were purchased from Sigma-Aldrich Dorset, UK and were of analytical grade unless stated otherwise.
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It appears that as the foulant blocks access to the pore entrances, the available surface area where protein can diffuse freely into the particle therefore decreases, which introduces increased resistance to mass transfer. Introduction Fouling of chromatographic resin over operational lifetimes can be a serious problem associated with industrial separations. Weak partitioning chromatography for anion exchange purification of monoclonal antibodies.